- In this lab, you will determine, through observation, which protease is secreted into the stomach, and which is secreted into the small intestine.
SAFETY: Wear safety goggles throughout this experiment. Acids and bases can damage eyes. If you get chemicals on your skin, wash with plenty of water.
MATERIALS:
Safety goggles
Wax pencil
6 test tubes
Test-tube rack
*18 cubes of boiled egg white (protein)
Metric ruler
10 mL graduated cylinder
10 mL of distilled water
**15 mL of dilute hydrochloric acid (HCl)
***15 mL of dilute sodium hydroxide (NaOH)
****20 mL of pepsin
*****20 mL of trypsin
Test-tube holder
Water bath or incubator at 37 degrees CPREPARATION OF SOLUTIONS AND EGGS:
* To prepare egg white (albumen), boil eggs for at least 5 minutes. Cool and cut the egg whites into 5mm X 5mm X 5mm cubes. One egg will make about 50 cubes.
** To prepare dilute (0.01 M) HCl, which has a pH of 2, add 50 mL of 0.1 M HCl to 300 mL of distilled water, then dilute to a final volume of 500 mL.
*** To prepare dilute (0.001 M) NaOH, which has a pH of 8, add 5 mL of 0.1 M NaOH to 300 mL of distilled water, then dilute to a final volume of 500 mL.
**** To prepare a 2% pepsin solution, place 4 g of dry pepsin in a flask and dilute it with 200 mL of distilled water. Use dilute HCL to adjust the pH of the solution to 2.
***** To prepare a 5% trypsin solution, place 10 g of dry pancreatin (a mixture of trypsin, amylopsin, and lipase) in a flask and dilute it with 200 mL distilled water. Use dilute NaOH to adjust the pH of the solution to 8.
PROCEDURE:
1. Use a wax pencil to label your test tubes directly on the glass. If they have white squares for labelling, use only a lead pencil on the white label.
Label the 6 test tubes as follows: C-2, C-8, P-2, P-8, T-2, and T-8.
- Use the test tubes marked C for your controls. the letter P indicates pepsin, and the letter T indicates trypsin. The number will indicate the pH of the test tube’s contents.
2. Place three cubes of boiled egg white in each test tube. Observe the size and appearance of the food cubes in each tube. Record your observations in your DATA TABLE.
3. Add 5 mL of distilled water to each test tube labelled C. Add 5 mL of pepsin to each test tube labelled P. Add 5 mL of trypsin to each test tube labelled T.
4. Add 5 mL of dilute hydrochloric acid (HCl) to each tube labelled 2. Add 5 mL of dilute sodium hydroxide (NaOH) to each tube labelled 8.
5. Place the test tubes in a water bath or incubator at 37 degrees C, and leave them there overnight. Make sure the temperature remains between 35 degrees C and 39 degrees C.
6. The next day, observe the tubes and note any changes in the size and appearance of the cubes of egg in each tube. Record your observations in your DATA TABLE.
DATA TABLE: Test
tubeBefore After 
Food sizeFood appearanceFood sizeFood appearanceC-2 C-8 P-2 P-8 T-2 T-8 QUESTIONS:
Q1. How did the contents of the tubes with a protease added differ from the tubes with no protease (control)?
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Q2. At what pH did pepsin break down protein most completely? _____
At what pH did trypsin break down protein most completely? _____
Q3. According to your results, what specific protease would break down protein in the small intestine? Justify your answer.
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Q4. According to your results, what specific protease would break down protein in the stomach? Justify your answer.
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Q5. What is the significance of using a water bath at 37 degrees C?
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Back to Lab Dad's Laboratory
- Use the test tubes marked C for your controls. the letter P indicates pepsin, and the letter T indicates trypsin. The number will indicate the pH of the test tube’s contents.
FINDING THE OPTIMUM pH
FOR TWO PROTEASE ENZYMES
Pepsin and Trypsin
INTRODUCTION:
During digestion, food is broken down in stages by a series of chemical reactions. These chemical reactions are catalyzed by digestive enzymes. A protease is an enzyme that breaks down proteins. Two proteases pepsin and trypsin, are secreted at different stages and at different sites during digestion. Each digestive enzyme works best at an optimum pH. The stomach has a pH of about 2, while the small intestine has a pH of between 7 and 8.